高粱拟盘多毛孢叶斑病病原菌鉴定、生物学特性及其防治药剂筛选

Identification and biological characterization of the pathogen causing Pestalotiopsis leaf spot in sorghum and screening for effective fungicides

摘要:

【背景】 随着山西省高粱种植面积逐年扩大，叶斑病发生普遍，危害加重且种类更趋复杂。【目的】 明确山西省高粱拟盘多毛孢叶斑病病原菌的种类及生物学特性，筛选有效防治该病害的化学药剂，为高粱拟盘多毛孢叶斑病的识别与防治提供理论依据。【方法】 采用组织分离法分离纯化病原菌，依据柯赫氏法则确认病原菌，结合形态学与分子生物学方法鉴定其种类，利用菌丝生长速率法与稀释平板法测定病原菌的生物学特性及其对杀菌剂的敏感性。【结果】 病原菌的形态学特征与肯尼亚拟盘多毛孢(Pestalotiopsis kenyana)一致，其ITS、TEF1-α和TUB基因序列与P. kenyana标准菌株CBS442.67的相似性均高于99%，基于供试菌株ITS-TEF1-α-TUB基因序列构建的系统发育树将其与P. kenyana聚在同一分支。病原菌在20-25 ℃、pH值为5.0-7.0、全黑暗，以葡萄糖为碳源，蛋白胨为氮源的培养基上生长最快。供试8种杀菌剂中，98%咯菌腈(0.105 4 mg/L)、98%吡唑醚菌酯(0.121 4 mg/L)、98%嘧菌酯(0.174 1 mg/L)、98%氟环唑(0.419 4 mg/L)和98%戊唑醇(0.924 9 mg/L)对菌株L-GLYK1的菌丝生长具有较好的毒力作用，EC₅₀值均小于1.0 mg/L。其中98%吡唑醚菌酯对菌株L-GLYK1的孢子萌发的抑制效果最好，EC₅₀为0.010 6 mg/L，98%戊唑醇(8.617 5 mg/L)与98%氟环唑(7.527 2 mg/L)也有较好的抑制效果。【结论】 肯尼亚拟盘多毛孢(Pestalotiopsis kenyana)为山西省高粱拟盘多毛孢叶斑病病原菌，明确了该病原菌的最佳培养条件，98%吡唑醚菌酯、98%戊唑醇与98%氟环唑可用于该病害药剂防治的进一步试验研究。

Abstract:

[Background] With the expansion of sorghum planting area in Shanxi Province, the occurrence of leaf spot becomes common, with aggravated harm and increasingly complex pathogen species. [Objective] To clarify the species and biological characteristics of the pathogen causing Pestalotiopsis leaf spot in sorghum in Shanxi Province, screen out effective fungicides that can effectively prevent and control the disease, and provide a theoretical basis for the identification and prevention of this disease in sorghum. [Methods] We employed the tissue culture method to isolate the pathogen and then confirmed the pathogen according to Koch's postulates. We identified the pathogen by combining morphological characteristics and molecular biological evidence, and adopted the mycelial growth method and dilution plate method to determine the biological characteristics and fungicide sensitivity of the pathogen. [Results] The morphological characteristics of the pathogen were consistent with those of Pestalotiopsis kenyana, and the ITS, TEF1-α, and TUB gene sequences of the pathogen showed the similarity above 99% compared with those of the P. kenyana standard strain CBS442.67. The phylogenetic tree constructed based on the ITS-TEF1-α-TUB gene sequences showed that the strain was in the same branch with P. kenyana. The suitable culture conditions for mycelial growth of the pathogen were 20-25 ℃, 24 h dark, pH 5.0-7.0, glucose as the carbon source, and peptone as the nitrogen source. Among the eight fungicides tested, 98% fludioxonil (0.105 4 mg/L), 98% pyraclostrobin (0.121 4 mg/L), 98% azoxystrobin (0.174 1 mg/L), 98% epoxiconazole (0.419 4 mg/L), and 98% tebuconazole (0.924 9 mg/L) showed stronger virulence on the mycelial growth of the pathogen strain L-GLYK1, with EC₅₀ values all less than 1.0 mg/L. Among them, 98% pyraclostrobin showed the best inhibitory effect on spore germination of strain L-GLYK1, with an EC₅₀ value of 0.010 6 mg/L. In addition, 98% tebuconazole (8.617 5 mg/L) and 98% epoxiconazole (7.527 2 mg/L) also exhibited good inhibitory effects. [Conclusion] P. kenyana is the pathogen causing Pestalotiopsis leaf spot in sorghum in Shanxi Province. The optimal culture conditions of this pathogen are determined, and 98% pyraclostrobin, 98% tebuconazole, and 98% epoxiconazole can be used for further experimental research on the prevention and control of this disease.